Regulation of ornithine decarboxylase activity by spermidine and the spermidine analogue N1N8-bis(ethyl)spermidine.

Polyamine biosynthesis in intact cells can be exquisitely controlled with exogenous polyamines through the regulation of rate-limiting biosynthetic enzymes, particularly ornithine decarboxylase (ODC). In an attempt to exploit this phenomenon as an antiproliferative strategy, certain polyamine analogues have been identified [Porter, Cavanaugh, Stolowich, Ganis, Kelly & Bergeron (1985) Cancer Res. 45, 2050-2057] which lower ODC activity in intact cells, have no direct inhibitory effects on ODC, are incapable of substituting for spermidine (SPD) in supporting cell growth, and are growth-inhibitory at micromolar concentrations. In the present study, the most effective of these analogues, N1N8-bis(ethyl)SPD (BES), is compared with SPD in its ability to regulate ODC activity in intact L1210 cells and in the mechanism(s) by which this is accomplished. With respect to time and dose-dependence of ODC suppression, both polyamines closely paralleled one another in their response curves, although BES was slightly less effective than SPD. Conditions of minimal treatment leading to near-maximal ODC suppression (70-80%) were determined and found to be 3 microM for 2 h with either SPD or BES. After such treatment, ODC activity was fully recovered within 2-4 h when cells were re-seeded in drug-free media. By assessing BES or [3H]SPD concentrations in treated and recovered cells, it was possible to deduce that an intracellular accumulation of BES or SPD equivalent to less than 6.5% of the combined cellular polyamine pool was sufficient to invoke ODC regulatory mechanisms. Decreases in ODC activity after BES or SPD treatment were closely paralleled by concomitant decreases in ODC protein. Since cellular ODC mRNA was not similarly decreased by either BES or SPD, it was concluded that translational and/or post-translational mechanisms, such as increased degradation of ODC protein or decreased translation of ODC mRNA, were probably responsible for regulation of enzyme activity. Experimental evidence indicated that neither of these mechanisms seemed to be mediated by cyclic AMP or ODC-antizyme induction. On the basis of the consistent similarities between BES and SPD in all parameters studied, it is concluded that the analogue most probably acts by the same mechanisms as SPD in regulating polyamine biosynthesis.     

Detecting Concealed Knowledge Using a Novel Attentional Blink Paradigm

Traditional concealed information paradigms rely on the idea that stimuli that are meaningful to a person (critical items) will draw attentional resources disproportionately, relative to stimuli that are not (irrelevant items), generating detectable differences on a suitable dependent variable (behavioral, psychophysiological, or neural). Here, we introduce a behavioral paradigm that could be used to reveal concealed information by exploiting the link between concealed information and attentional processes more directly. This novel paradigm is based on the attentional blink phenomenon in which detection of a stimulus reduces detection accuracy rates of subsequent target stimuli within a 200–500 ms time window. We hypothesized that a well-known face used as a critical item could capture attentional resources automatically, making it harder to detect the occurrence of a subsequently presented target face. The results confirmed this hypothesis, and showed that concealed knowledge of a famous face could be detected in 9 out of 12 individuals by looking for a relative dip in target detection accuracy after the presentation of a critical item.     

Characterization of human spermidine/spermine N1-acetyltransferase purified from cultured melanoma cells

Extreme inducibility of spermidine/spermine acetyltransferase (SSAT) by bis-ethyl derivatives of spermine in human large cell lung carcinoma and melanoma cells has prompted biochemical characterization of the purified enzyme. Treatment of human MALME-3 melanoma cells with 10 microM N1,N11-bis(ethyl)norspermine (BENSPM) for 48-72 h increased SSAT activity by some 1000- to 4000-fold and enabled purification of the enzyme by established procedures--binding on immobilized spermine and elution with spermine followed by binding on Matrex Blue A and elution with coenzyme A. The enzyme showed a single band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a single subunit species and molecular weight of approximately 20,300 Da. By gel permeation chromatography, the holoenzyme was found to have a molecular weight of 80,000 Da, suggesting a total of four identical subunits. Purified SSAT had a specific activity of 285 mumol/min/mg for spermidine and Km values of 5.9 microM for acetylcoenzyme A, 55 microM for spermidine, 5 microM for spermine, 36 microM for N1-acetylspermine, 1.6 microM for norspermidine, and 4 microM for norspermine. Homologs of BENSPM were found to be competitive inhibitors of spermidine acetylation, with Ki values of 0.8 microM for BENSPM, 1.9 microM for N1,N12-bis-(ethyl)spermine and 17 microM for N1,N14-bis-(ethyl)-homospermine. Correlation of these values with the relative abilities of the homologs to increase SSAT in intact cells suggests that formation of an enzyme inhibitor complex may play a contributing role in enzyme induction.     

Pedodiversity deserves attention in plant biodiversity research

We suggest that pedodiversity, an expression of environmental heterogeneity, should deserve the attention of plant ecologists interested to study the spatial pattern of plant biodiversity at different scales. Using the FAO pedological and the IUCN-WCMC biological data bases of the world countries, we show that the prediction on plant biodiversity (number of vascular plants) improves significantly when the extent of the area is combined with pedodiversity in a multiple polynomial regression. Partial correlation analysis proves that, by removing the effect of pedodiversity, the correlation “number of species–area” remains statistically significant for the tropical countries while it loses significance for the countries outside the tropics.     

A comparison of root characteristics in relation to nutrient and water stress in two maize hybrids

Root responses of maize (Zea mays L.) to limited nutrients and water availability were evaluated in two highly productive full-season hybrids, DK585 and Santos (Dekalb – Monsanto), in laboratory, pot and field tests. In the laboratory, under optimal nutrient and water supply, seedlings of DK585 had higher growth (leaves and roots). Under nitrate or sulphate deprivation, DK585 showed better ability in adapting its root/shoot ratio to stress conditions, whereas Santos showed less plastic behaviour. This morphological trait of DK585 was associated with higher sulphate and constitutive nitrate influxes. In pot trials (plants with four to five leaves), DK585 maintained a high transpiration level to very low values (around 0.2) of FTSW (fraction of transpirable soil water), whereas Santos showed a higher response to soil drying. The latter reduced the rate of transpiration starting from a FTSW of about 0.6. In the open field (trial in 2000, Legnaro, NE Italy), in conditions of fluctuating combined water and nitrogen stress, DK585 at flowering reached greater root length density (RLD) than Santos in deep layers (50–100-cm interval depth) of positions further from the plant. However, in these conditions, the yield of DK585 was found to be only slightly higher than that of Santos (8.88 vs. 8.49 t ha^–1 d.w.). An overall evaluation of the two hybrids indicates the more conservative strategy towards limited water and nutrient resources in Santos, and a greater tendency towards stress avoidance in DK585.     

Hydrocortisone and progesterone regulation of the proliferation,morphogenesis, and functional differentiation of normal rat mammary epithelial cells in three dimensional primary culture

The mechanisms of action of, and resistance to, the steroidal regulators of normal mammary epithelial and breast cancer cell development are only partially understood. A major obstacle to research progress has been the difficulty in supporting physiologically relevant development of normal mammary epithelial cells (MEC) under defined serum-free conditions. A primary culture system was developed in our laboratory that permits nonfunctional rat MEC to undergo extensive proliferation, functional differentiation, as well as multilobular and lobuloductal branching alveolar morphogenesis. In the studies reported here, the contributions of hydrocortisone and progesterone during the coordinate induction of cellular proliferation, organoid morphogenesis, and functional capacity were assessed. Hydrocortisone (0.1–10 μg/ml) induced alveolar and multilobular branching morphogenesis, suppressed lobuloductal branching morphogenesis, and enhanced casein accumulation. Hydrocortisone also played a role in maintaining alveolar as well as multilobular branching morphogenesis and casein levels. Progesterone (0.01–1 μg/ml) induced cellular proliferation as well as multilobular and lobuloductal branching morphogenesis, and suppressed casein accumulation. At a supraphysiological concentration (10 μg/ml), progesterone inhibited cell growth, alveolar branching morphogenesis, and casein accumulation. MEC cultured without progesterone for up to 1 week retained the ability to respond when subsequently exposed to this steroid. Reversibility studies suggested that progesterone was required for the induction, but not the maintenance of the mitogenic, morphogenic, and lactogenic effects. This physiologically relevant primary culture system can be used to study the factors that regulate steroid responsiveness as well as the cross-talk between steroid and growth factor receptor signaling pathways in normal MEC and breast cancer cells. © 1995 Wiley-Liss, Inc.